Adenanthera pavonina L, Fabaceae alt. Leguminosae, Bengali name 'rakta kombol', is an Indian medicinal plant. It is endemic to Southern China and India, and widely naturalized in Malaysia, Western and Eastern Africa as well as in most islands of both the Pacific and Caribbean regions. This plant has been used in traditional medicine for the treatment of asthma, boil, diarrhoea, gout, inflammations, rheumatism, tumour and ulcers, and as a tonic. The dried and ground bark of A. pavonina L. was extracted, successively, with petroleum ether (PE), dichloromethane (DCM), ethyl acetate (EtOAc) and methanol (MeOH), and the resulting extracts were assessed in vivo for anti-inflammatory activity on carrageenan-induced rat hind paw oedema as a model of inflammation. The extracts were administered orally at the doses of 200 and 400 mg/kg body weight, and statistically significant (p<0.001) anti-inflammatory effects were observed in a dose dependant manner. The MeOH extract (400 mg/kg) showed 37.10% (p<0.01) inhibition of inflammation at the first hour of the study and, the DCM extract (400 mg/kg) exhibited 33.11% (p<0.001) inhibition of inflammation at the third hour of the study which was comparable with that of reference standard drug dichlofenac sodium. The results of this study supported some of the traditional medicinal uses of this plant.

Adenanthera pavonina L., Fabaceae alt. Leguminosae, commonly known in Bangladesh as 'rakta kombol', is an important medicinal plant from the 'Indian subcontinent'. This species is endemic to Southern China and India, and widely naturalized in Malaysia, Western and Eastern Africa as well as in most islands of both the Pacific and Caribbean regions (Balogun & Fetuga, 1985; GRIN Databases, 2009). While the wood of this plant has been used for decorative wood products, the fruits are consumed by local people (Benthall, 1946; Clark & Thaman, 1993). Various parts of this plant have also been used in traditional medicine for the treatment of asthma, boil, diarrhoea, gout, inflammations, rheumatism, tumour and ulcers, and as a tonic (Watt & Breyer-Brandwijk, 1962; Kirtikar & Basu, 1981; Burkil, 1994; Dr Duke's, 2009). The seeds of this plant have been found to be effective in treating cardiovascular diseases during pregnancy. Previous phytochemical studies on this plant revealed the presence of various secondary metabolites including mainly flavonoids, steroids, saponins and triterpenoids (Chandra et al., 1982; Yeoh et al., 1984). The antibacterial, antifungal, antioxidant, cytotoxic and blood pressure reducing activities of the extracts of A. pavonina have recently been reported (Jayasinghe et al., 2006; Rodrigo et al., 2007; Adedapo et al., 2009). We now report on the in vivo anti-inflammatory activity of various extracts of the bark of A. pavonina in carragennan-induced rat oedema model.

The barks of Adenanthera pavonina L., Fabaceae, were collected from a mature plant from the Jahangirnagar University campus, Savar, Dhaka, Bangladesh, in March 2008. A voucher specimen representing this collection (accession No.-DACB34196) was deposited in the National Herbarium of Bangladesh.

Extraction

The barks of A. pavonina were shade-dried, pulverized by a mechanical grinder and passed through a 60 mesh sieve to obtain fine powder, a portion (1.0 kg) of which was extracted, successively, with PE, DCM, EtOAc and MeOH (2 L each). All extracts were evaporated to dryness under reduced pressure. The yields of the PE, DCM, EtOAc and MeOH extracts were 2.1 g, 3.7 g, 2.5 g and 30.0 g, respectively.

Animals

Long Evan rats of either sex (weighing 130-180 g) were collected from the International Centre for Diarrhoeal Diseases and Research, Bangladesh (ICDDR, B). The animals were kept in polyvinyl cages under controlled room temperature (25±2 ºC) in the laboratory environment under 12 h dark and 12 h light cycle for seven days. The ICDDR, B formulated food pellets were supplied to the animals and water at libitum. Animals were fasted overnight and weighed before the experiment. The study involving rats was approved by the Ethical Review Committee (approval number: SCB090221) Biological Science, University of Dhaka, Bangladesh, and the experiments were carried out strictly in accordance with the guidelines provided by the World Health Organization.

The anti-inflammatory activity of the extracts of A. pavonina was measured by the carrageenan-induced rat paw oedema model (Winter et al., 1962). Experimental animals were randomly selected, irrespective of sexes, and divided into ten groups consisting of 5 rats in each group. The test compounds were administered orally to each group at 200 and 400 mg/kg body weight. After 1 h of treatment, acute inflammation was produced by sub-planter injection of 0.1 mL of 1% suspension of carrageenan in normal saline in the right hind paw of the rats. Another animal group was treated with dichlofenac sodium at a dose of 15 mg/kg per orally (p.o.) as a reference standard. The saline treated group was considered as control. The paw volume was measured plethysmometrically (Ugo Basile, Italy) at 1, 2, 3 and 4 h after the carrageenan injection. Results were expressed as percentage of inhibition of oedema calculated by the formula- (1 - Vt/Vc) × 100, where Vt and Vc are the mean paw volume in the treated and controlled groups, respectively.

Statistical analysis

Statistical analysis was performed using SPSS-11.5 statistical Software for Windows. Experimental values were expressed as mean±SEM. Independent Sample t-test was carried out for statistical comparison. Statistical significance was considered to be indicated by a p value<0.05 in all cases.

RESULTS AND DISCUSSION

The extracts of the barks of A. pavonina were evaluated for anti-inflammatory activity using the carrageenan-induced rat paw oedema model at dose levels of 200 and 400 mg/kg body weight, where dichlofenac sodium was used as a positive reference standard (Table 1). The results showed that the PE fraction exhibited 25.6% and 27.8% inhibition of the paw oedema at the doses of 200 and 400 mg/kg, respectively at the first hour of the study. The DCM fraction exhibited maximum inhibition at the third hour of the study with 31.61% and 33.11% inhibition at the doses of 200 and 400 mg/kg, respectively. The DCM extract exhibited significant (p<0.001) inhibition of paw oedema at different time intervals. The EtOAc extraction also demonstrated the maximum inhibition (28.63%) after the first hour of the study at the dose of 400 mg/kg body weight only. On the other hand the MeOH extract displayed maximum anti-inflammatory effect at the first hour with 30.85% and 37.1% inhibition at the doses of 200 mg/kg and 400 mg/kg body weight, respectively, but at the second and third hour, the effects were observed in decreasing pattern. Among the extracts, the DCM extract showed the maximum effect at the third hour of the experiment, whereas the MeOH extracts exhibited the maximum effect at the first hour of the study at 400 mg/kg. Both these extracts exhibited the anti-inflammatory effect in a dose dependent manner.

The carrageenan induced paw oedema model is known to be sensitive to cyclooxygenase inhibition which has been used to evaluate the effect of non-steroidal anti-inflammatory agents (Seibert et al., 1994; Appleton et al., 1995). The time response of oedema in rat is believed to be biphasic (Vinegar et al., 1969). The first phase which occurs between 0.5 to 2 h after injection of the fluogestic agent has been attributed to the release of histamine or serotonin. The second phase of the inflammatory reaction is caused by the release of bradykinin, protease, prostaglandin and lysosome (Crunkhorn & Meacock, 1971; Di Rosa & Willoughby, 1971). Therefore it can be assumed that the inhibitory effect of the MeOH extract on carrageenan induced inflammation could be due to the inhibition of histamine or serotonin. On the other hand, the maximum inhibitory activity of the DCM extract at the third hour of the study was due to the inhibition of the enzyme cyclooxygenase leading to the inhibition of prostaglandin synthesis.

The significant in vivo anti-inflammatory activities exhibited by the extracts of A. pavonina in the carrageenan-induced rat paw oedema model provide some scientific evidence in support of the traditional uses of this plant for treatment of various inflammatory conditions, e.g. gout, rheumatism and tumours.

ACKNOWLEDGEMENTS

The authors are thankful to Prof. Mohammad Rahmatullah, Pro-Vice Chancellor, University of Development Alternative, Dhaka, Bangladesh, for provision of the facilities for biological experiments. Arzumand Ara is grateful to the University Grant Commission (UGC), Bangladesh, for the award of a fellowship.